Use of polyunsaturated fatty acids for the primary prevention of major cardiovascular events

ABSTRACT

The use of polyunsaturated fatty acids of the ω-3 series such as eicosapentaenoic acid (EPA, C 20:5  ω-3), docosahexaenoic acid (DHA, C 22:6  ω-3), or their pharmaceutically acceptable derivatives is described for the primary prevention of major cardiovascular events in subjects who have not undergone previous infarct episodes.

CROSS-REFERENCE TO RELATED APPLICATION

This Application is a Divisional, which claims the benefit of pendingU.S. patent application Ser. No. 10/984,485, filed on Nov. 19, 2007,which is a divisional of U.S. patent application Ser. No. 10/778,182,filed Feb. 17, 2004, issued as U.S. Pat. No. 7,553,870, on Jun. 30,2009, which in turn is a Divisional of Ser. No. 10/281,208, filed Oct.28, 2002, which is now abandoned. The disclosures of the priorapplication are hereby incorporated herein in their entirety byreference.

The invention relates to the use of polyunsaturated fatty acids for theprimary prevention of major cardiovascular events.

In particular, the invention concerns the use of polyunsaturated fattyacids of the ω-3 series such as eicosapentaenoic acid (EPA, C_(20:5)ω-3) docosahexaenoic acid (DHA, C_(22:6) ω-3), or their pharmaceuticallyacceptable derivatives, either alone or mixed together, for the primaryprevention of major cardiovascular events.

The beneficial effects of polyunsaturated fatty acids of the ω-3 serieson multiple risk factors for cardiovascular illnesses are well known;for example the patents IT 1235879, U.S. Pat. No. 5,502,077, U.S. Pat.No. 5,656,667 and U.S. Pat. No. 5,698,594 refer respectively tohypertriglyceridemia, defects of the cholesterol level and hypertension.However, each of the cited documents deal with the treatment of riskfactors, not with real and proclaimed illnesses.

U.S. Pat. No. 5,753,703 describes the use of L-carnitine or itsderivatives in association with polyunsaturated fatty acids of the ω-3series or their esters, in particular EPA and DHA, for the preventionand treatment of cardiovascular disorders, vascular pathologies,diabetic peripheral neuropathies, and atherosclerotic, thromboembolyticand tissue disorders.

EP-B-0409903 describes a process for preparing high concentrationmixtures of EPA and DHA and/or their esters useful for treatinghyperlipemia and related pathologies, thrombosis, cardiac infarct,platelet aggregation, as anticoagulants in the prevention ofatherosclerosis, for the treatment of cerebral infarct, of lesions andocclusions caused by vasomotor spasms, of diabetes and itscomplications, of chronic and acute inflammations, of autoimmunesymptoms, in the prevention of side effects caused by non-steroidanti-inflammatories at the gastrointestinal level and in tumourprevention.

CN 1082909 describes compositions based on ethyl esters of EPA and DHAand other polyunsaturated fatty acids of the ω-3 series in associationwith soya phospholipids, oenothera odorata and ginkgetin, asantithrombotic and antidementia agents for treating for example dementiaand infarct of the myocardium.

U.S. Pat. No. 5,760,081 describes a method for preventing imminentfibrillation of the myocardial ventricle by intravenous infusion of acomposition containing EPA, where the subject at risk of imminentfibrillation has already often been the protagonist of an episode ofinfarct of the myocardium and where the infusion is effected within 3hours of the infarct episode, possibly using intracardiac injection.These are always situations of extreme emergency and of parenteralintervention, for the specific treatment of ventricular fibrillation.

Swann et al., Clinical Drug Investigation 15 (6), 473, 1998 have alsoshown that the administration of EPA and DHA ethylesters, at a dose of 4g per day, leads to a decrease in triglycerides and total apolipoproteinC III and to an increase in antithrombin III, in subjects with abnormalplasmatic lipoprotein symptoms and have undergone an infarct of themyocardium, they having consequently suggested that an administration ofthese compositions can result in an improvement in the lipoprotein leveland hence a decrease in the relative risk factors.

WO 00/48592 describes the use of a mixture of EPA and DHA ethylesters inquantities greater than 25 wt. % for preventing death, in particular“sudden death” in patients who have already suffered an infarct of themyocardium.

This therefore represents the use of said mixture in so-called secondarydeath prevention, i.e. in subjects who have already suffered infarct.

The prevention of cardiovascular damage by means of fatty acid mixturesdescribed in the state of the art is therefore focused on “secondary”prevention of cardiovascular damage. i.e. aimed at protecting a subjectwho has already suffered an infarct, whereas “primary” prevention ofmajor cardiovascular events, i.e. prevention in subjects who, whileaffected by various pathologies of the cardiocirculatory and/orcardiorespiratory systems, have not yet suffered an infarct episode,constitutes a technical problem which is still felt in this sector.

According to a first aspect the invention relates to the use ofpolyunsaturated fatty acids of the ω-3 series for the preparation of adrug useful in the primary prevention of a major cardiovascular event insubjects who have not undergone previous infarct episodes, wherein thefatty acids comprise eicosapentaenoic acid (EPA) and/or docosahexaenoicacid (DHA) and/or at least one pharmaceutically acceptable derivativethereof, in quantities greater than or equal to 25 wt % on the totalfatty acid weight.

In the present description, the expression “polyunsaturated fatty acidsof the ω-3 series” means those long-chain polyunsaturated fatty acids,generally C₁₆-C₂₄, containing fish oils, in particular those having aC₂₀-C₂₂ chain, which are predominant in purification processes.

The expression “major cardiovascular event” means in particular thoseevents which involve reversible or irreversible cardiovascular damage,such as infarct of the myocardium and of individual coronary branches,death from cardiac causes, sudden death, etc., besides to infarct,broadly speaking, ictus etc., and those conditions prodromal to suchmajor events, such as myocardial fibrillation, atrial and/or ventricularfibrillation, etc. Said major cardiovascular events are usually inducedby various cardiocirculatory and cardiorespiratory pathologies such ascoronary ischemic illness not displayed by previous infarct episodes,and by serious hypoxic/anoxic states caused by a sudden lack of oxygen(for example during anesthesia, surgery, etc.), possibly in the presenceof conditions which contemplate an increase in the oxygen requirement.(accentuated physical stress, drug abuse, acute hypertensive crises,etc.) and analogous acute and chronic pathologies due to cardiac defectsof electrical and/or mechanical type.

The subjects affected by pathologies of the cardiocirculatory andcardiorespiratory system, hence not simply at prospective risk due tohypertriglyceridemia, hypertension or other, are representative ofsubjects definable at various levels as cardiopaths, by being affected,for example, by coronary ischemia detectable by coronarography,scintigraphy of the myocardium, electrocardiogram (ECG) under stress,etc., against which interventions of revascularization (angioplasty) orother possible pharmacological or invasive treatments have beenproposed, and of subjects affected by electrical hyperexcitability ofthe myocardium cells, disorder of the diffusion of electrical excitementor of electrical conduction (arrhythmia, fibrillation, etc.) or by otherdefects of mechanical type (cardiac insufficiency, decompensation),possibly aggravated by concomitant pathologies such as diabetes.

The use of polyunsaturated fatty acids of the ω-3 series according tothe invention is particularly indicated if the occurrence of a majorevent is predicted, such as an infarct, in particular of the myocardium,death from a cardiological cause, or sudden death, and where such anoccurrence takes place in cardiopathic subjects affected, for example,by coronary ischemia, arrhythmia, atrial and/or ventricularfibrillation, electrical hyperexcitability of the myocardium cells,disorder of the diffusion of electrical excitement or of electricalconduction of the myocardium, or cardiac disorders of mechanical type,for example cardiac insufficiency or cardiac decompensation, possiblyaffected by diabetic pathology concomitant with the cardiopathy.

Preferably, the content of EPA and/or DHA and/or of the at least onederivative thereof is between 50% and 100%, in particular between 75%and 95%, and more preferably about 85% by weight on the total fatty acidweight.

The preferred EPA and/or DHA derivatives are selected from thecorresponding C₁-C₃ alkyl esters and/or from their salts withpharmaceutically acceptable bases such as sodium hydroxide, lysine,arginine or aminoalcohols such as choline. The ethylesters of EPA andDHA, in particular mixed together in any concentration and percentage,are the most preferred.

The drug is administered preferably orally, in particular in the form ofsoft gelatin capsules. For oral use, the unit dose generally comprises100-1000 mg of polyunsaturated fatty acids of the ω-3 series, preferably500-1000 mg or 300-500 mg, the total dose being usually around 0.1-3.0 gper day or per alternate day, according to the case concerned, andpreferably 0.3-2.0 g per day and in particular 1.0 g per day. Theeffective dose of the drug suitable for the use of the invention is1.0-60.0 mg/kg of body weight/day.

Other types of formulation for oral administration are also suitable forthe purposes of the invention; for example hard capsules or tablets, inwhich the polyunsaturated fatty acids are adsorbed on solid supports. Itis also possible to use emulsions, granulates in dispersing excipients,syrups, droplets, etc., and other forms of administration able to ensuresystemic absorption of the drug, such as sterile solutions or emulsionsand the like, suitable for parenteral use and the like, as evaluated bythe expert of the art, on the basis of the severity of the pathology.

Those compositions illustrated in the European Pharmacopea 2000 (EuPh.2000), containing quantities greater than or equal to 80 wt % ofmixtures of EPA and DHA ethylesters and a total of ω-3 polyunsaturatedfatty acid ethylesters greater than or equal to 90 wt % are alsosuitable for the purposes of the present invention.

The aforestated compositions and the drugs suitable for the use of theinvention can be prepared by methods known to the expert of the art,such as those described in U.S. Pat. No. 5,130,061, WO 89/11521, IT1235879, JP 02/25447, which are incorporated into the presentdescription with regard to the method of preparation.

The drug suitable for use according to the present invention can alsocomprise other active principles and/or drugs, in association,possessing activity complementary to or synergic with that of the drugsuitable for use according to the invention, and also at least onepharmaceutically acceptable vehicle and/or one diluent and/or onesurfactant and/or one thickener and/or one binder and/or one lubricantand/or one aromatizer and/or one colorant and/or one stabilizer and thelike, which can easily be selected by the expert of the art. Of thestabilizers, antioxidants such as vitamin E (tocopherol), ascorbylpalmitate, ascorbic acid, hydroxytoluene and the like, which can beeasily selected by the expert of the art, are particularly preferred.

According to another aspect, the invention relates to a method for theprimary prevention of a major cardiovascular event in subjects who havenot undergone previous infarct episodes, comprising the administrationof an effective dose of a drug comprising polyunsaturated fatty acids ofthe ω-3 series as hereinbefore described. In particular, the method ofthe invention is indicated whenever the occurrence of a major cardiacevent is predicted such as an infarct, in particular of the myocardium,death from a cardiological cause or sudden death.

The following examples illustrate the invention but without limiting it.

The compositions illustrated in the following table were prepared by themethods described in U.S. Pat. No. 5,130,061 (compositions A, C, D, F),IT 1235879 (composition B), JP 02/25447 (composition E) and WO 89/11521(compositions G-I).

All the quantities indicated in the following table express percentagesby weight on the total weight of polyunsaturated fatty acids of the ω-3series.

A¹ B¹ C¹ D¹ E¹ F¹ G¹ H² I³ EPA >40 >44 >40 >25 >80 >20 <15 >40 >40DHA >34 >30 >34 >20 <10 >25 >80 >30 >30 EPA +DHA >85 >80 >80 >50 >80 >50 >85 >80 >80 esters⁴ >3 — tot. esters⁵ >90α-tocopherol 0.03 0.03 0.1 0.3 0.1 0.3 0.03 0.1 0.1 ¹ethyl esters; ²freeacids; ³sodium salts; ⁴ethyl esters of other (C₂₀, C₂₁, C₂₂) ω-3 acids;⁵total ethyl esters of ω-3 acids.

EXAMPLE 2

The compositions illustrated in the following table, relative to softgelatin capsules containing 1 g of polyunsaturated fatty acid ethylesters, were prepared by methods known in the art.

A (mg) B (mg) C (mg) EPA¹ 525 — >400 DHA¹ 315 — >340 EPA + DHA¹ 850 >800Total ω-3¹ — >900 d-α-tocopherol 4 I.U. — 4 I.U. d,l-α-tocopherol 0.3gelatin 246 — 246 gelatin succinate 233 glycerol 118 67 118 OFR 2.27 —2.27 OFG 1.27 1.27 SOB 1.09 SPOB 0.54 ¹ethyl esters; OFR: red iron oxideOFG: yellow iron oxide; SOB: sodium p- oxybenzoate; SPOB: sodium propylp-oxybenzoate; I.U.: international units.Pharmacological Activity

The pharmacological activity of the compositions of the invention wasevaluated on the basis of tests carried out on small laboratory animals(mouse, guinea pig, rat); this experimental model was chosen because ofthe ability to make rapid and highly reproducible verifications and touse a sufficiently large number of animals, such as to enable astatistically accurate evaluation of the results to be made withoutexposing the patient to risk, with evident ethical implications.

During the course of these tests, groups of animals were pretreatedrepeatedly with the formulations of Examples 1 and 2 and then, incomparison with untreated groups, were subjected to the action ofcardiotoxic or respiration-depressive substances, then visuallymeasuring protection against death, or—by means of electrocardiographicrecording—measuring the delay in the start of initial cardiac arrhythmiaor of ventricular tachycardia and above all the delay in or theprevention of animal death due to sudden cardiac and/or respiratoryarrest.

Using an analogous experimental model, cardiological pathology, coronaryischemia and a state of infarct were induced by coronary ligatureinstead of by cardiotoxic agents.

Test 1

The experimental sudden death model was obtained by cardiac arrestinduced by intravenous (i.v.) administration of a cardiotoxic agent(ouabain). In preliminary tests, various doses of ouabain wereadministered to non-anesthetized guinea pigs of both sexes of weight300-380 g, in order to determine the minimum lethal dose for 100% of theanimals within 15 minutes from i.v. injection (240 mg/kg, intravenouslyadministered over 3 minutes).

Two groups of 20 guinea pigs were then treated with 50 and 100 mg/kg ofa composition containing 85% of EPA and DHA ethylesters (Ex. 1,composition A) for 10 days. After 2 hours from the last administrationthe two groups of guinea pigs and a further untreated group, used ascontrol, were treated with 240 mg/kg i.v. of ouabain, recordingmortality within the subsequent 15 minutes.

Results expressed as survivors after 15 minutes:

Controls 00/20  50 mg/kg 11/20 100 mg/kg 16/20Test 2

3 groups of 15 male mice, initial weight 25-32 g, were treated orallyfor 15 days with physiological solution (control group) and with 50 or100 mg/kg of a composition containing 85% of EPA and DHA ethylesters(Ex. 1, composition A).

60 minutes after the end of the last treatment, the animals of all thegroups were treated with sodium pentabarbital i.p. (50 mg/kg) and thenwith aconitine i.v. (0.25 mg/kg). The times of appearance of cardiacarrhythmia (deviation >5 seconds from the normal sinus rhythm), ofventricular fibrillation and of cardiac arrest were determined byelectrocardiograph recording. Results expressed as mean±standarddeviation (seconds) on the positive animals.

T t₁ (sec) t₂ (sec) t₃ (sec) S C 123 ± 12 174 ± 7  214 ± 32 00/15(15/15) (15/15) (15/15)  50 mg/kg 168 ± 8  235 ± 16 350 ± 26 09/15(08/15) (06/15) (06/15) 100 mg/kg 195 ± 15 284 ± 18 378 ± 35 12/15(05/15) (03/15) (03/15) T—treatment; t₁—time of appearance of arrhythmia(number of animals); t₂—time of appearance of fibrillation (number ofanimals); t₃—time of cardiac arrest (number of animals); S—survivorsafter 15 minutes; C—control (with physiological solution).Test 3

2 groups of 20 male rats, initial weight 310-350 g, were treated orallyfor 15 days with physiological solution (control group) and with 100mg/kg of a composition containing >80% of EPA and DHA ethylesters (Ex.1, composition B). The rats of the 2 groups were then anesthetized withsodium pentobarbital i.p. (50 mg/kg), then subjected to ligature of theleft anterior descending coronary artery, which allows blood flow to theleft ventricle, so inducing an acute ischemic state of the myocardium.During the subsequent 15 minutes the duration of ventricularfibrillation was recorded by ECG, this either resolving itselfspontaneously or concluding with sudden death.

Results

T F (sec) Mortality S (%) C 190 ± 24 16/20 20 (18/20) 100 mg/kg 55 ± 501/20 95 (04/20) T—treatment; F—duration of fibrillation; S—survivalsafter 15 minutes; C—control (with physiological solution).Test 4

The experimental model implemented for sudden death by respiratoryarrest involves its inducement by chloroform inhalation.

4 groups of 10 male mice, initial weight 26-32 g, were treated orallyfor 5 days with physiological solution (control group) and with 10, 30and 60 mg/kg of a composition containing 80% of EPA and DHA ethylesters(Ex. 1, composition C).

60 minutes after the end of the last treatment, the animals were exposedto chloroform until respiratory arrest had occurred. The animals werethen checked for tachyarrhythmia of the myocardium induced by thehypoxic state, this either resolving itself spontaneously within thenext 15 minutes or concluding with death of the animal.

Results

Protection from Treatment tachyarrhythmia Survivals Control 00/10 03/1010 mg/kg 04/10 06/10 30 mg/kg 07/10 08/10 60 mg/kg 09/10 10/10Test 5

2 groups of 20 male rats were treated as on Test 3, with physiologicalsolution and with the same EPA and DHA composition (Example 1,composition B).

Ligature of the circumflex coronary artery was then effected, withconsequent reduction in the contractile capacity of the myocardium andof the ejection fraction. The mortality of 18/20 animals of the controlgroup fell to 4/20 of the treated group, during the course of thesubsequent 60 minutes.

The clinical results of the tests demonstrate the pharmacologicalactivity of the polyunsaturated fatty acids of the ω-3 series in theprimary prevention of major cardiovascular events in subjects who havenot undergone previous infarct episodes.

What is claimed:
 1. A method of reducing the probability of anoccurrence of a major cardiovascular event selected from the groupconsisting of: infarct of the myocardium, death from a cardiac cause,and sudden death, in a subject who has not undergone a previous infarctepisode and who is affected by a condition selected from the groupconsisting of: cardiac insufficiency, and cardiac decompensation,comprising orally administering to the subject an effective amount of apharmaceutical composition comprising ω-3 fatty acid ethyl esters in anamount of at least 90% by weight of total fatty acids, wherein the ω-3fatty acid ethyl esters consist essentially of a mixture ofeicosapentaenoic acid (EPA) ethyl ester and/or docosahexaenoic acid(DHA) ethyl ester in an amount of at least 80% by weight of total fattyacids.
 2. The method of claim 1, wherein the effective amount is between1-60 mg/kg of the subject's body weight per day.
 3. The method of claim1, wherein the pharmaceutical composition is in an oral, unit dose formcomprising 500-1000 mg of the ω-3 fatty acids.